ABSTRACT HIV plays a complicated game of hide-and-seek with the human immune system and antiretroviral therapy (ART). The virus hides in various anatomic compartments, like male genital tract (MGT), which each differ in their vascular, physiochemical and immunological environments and ART penetration. These features shape how HIV adapts to these compartments, and may have considerable implications for the HIV cure strategies. The MGT is a unique HIV sanctuary and its accessibility makes it a prime candidate for investigating how HIV persists and propagates in anatomic tissues in the setting of modern ART. The goals of this proposal are to determine: 1) if the MGT, and prostate in particular, supports propagating HIV infection in the setting of modern ART, 2) if HIV traffics between blood and MGT during ART, and 3) how clinical and virologic factors influence residual replication in the MGT. To accomplish these goals, we will evaluate 30 HIV-infected men who started ART during acute infection and virologically suppressed for > 96 weeks and for whom one paired blood and semen sample are available prior to ART initiation. We will prospectively collect 3 timepoint sampling of paired blood, semen and urine: 2 samplings after prostate stimulation and 1 without prior prostate stimulation. All collected >6 months apart. From these samples, we will: measure low level HIV RNA levels, sequence full length env HIV DNA populations, and measure clinical and virological factors. With these data, we will characterize the factors associated with viral propagation (i.e. evolution) during ART and migration between blood and semen. Understanding the interplay between HIV and its local environment is a pre-requisite for designing HIV eradication strategies. 1